U.S. patent application number 14/062234 was filed with the patent office on 2014-04-24 for genome explorer system to process and present nucleotide variations in genome sequence data.
This patent application is currently assigned to Complete Genomics, Inc.. The applicant listed for this patent is Complete Genomics, Inc.. Invention is credited to Julie Adams, Mirko Buholzer.
Application Number | 20140115515 14/062234 |
Document ID | / |
Family ID | 50486557 |
Filed Date | 2014-04-24 |
United States Patent
Application |
20140115515 |
Kind Code |
A1 |
Adams; Julie ; et
al. |
April 24, 2014 |
Genome Explorer System to Process and Present Nucleotide Variations
in Genome Sequence Data
Abstract
This disclosure provides a technology for users to gain
first-hand knowledge and experience with interpreting whole
genomes. The technology graphically depicts variations in genome
sequences in an expandable display, and provides a platform whereby
the user may find and research the biological significance of such
variants. The technology also provides a unique collaborative
environment designed to capture and improve the collective
knowledge of the participating community.
Inventors: |
Adams; Julie; (Pleasanton,
CA) ; Buholzer; Mirko; (Cupertino, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Complete Genomics, Inc. |
Mountain View |
CA |
US |
|
|
Assignee: |
Complete Genomics, Inc.
Mountain View
CA
|
Family ID: |
50486557 |
Appl. No.: |
14/062234 |
Filed: |
October 24, 2013 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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61717773 |
Oct 24, 2012 |
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Current U.S.
Class: |
715/771 |
Current CPC
Class: |
G16B 30/00 20190201;
G16B 20/00 20190201; G16B 45/00 20190201; G16B 50/00 20190201 |
Class at
Publication: |
715/771 |
International
Class: |
G06F 19/22 20060101
G06F019/22 |
Claims
1. A method for displaying variations between complex sequences,
the method comprising: (a) receiving a sample complex sequence for
a test sample; (b) aligning, with a computer system, the sample
complex sequence with a reference complex sequence for a reference
sample; (c) identifying, with the computer system, a plurality of
variations between the sample complex sequence and the reference
complex sequence; (d) providing a user interface for controlling a
display of a comparative gene sequence map, the display showing
where the plurality of variations are located; (e) receiving, at
the user interface, a selection of a region within the genetic
sequence map, the region being less than half the entire
comparative gene sequence map; and (f) reconfiguring the display of
the comparative gene sequence map such that the region is
magnified.
2. The method of claim 1, wherein the region is magnified in
proportion to the size of the region.
3. The method of claim 1, wherein the region is magnified at least
two times.
4. The method of claim 1, wherein the region is magnified such that
the region is presented across at least about half of the width of
the display.
5. The method of any preceding claim, further comprising:
determining whether the region is smaller than a specified size; if
the region is smaller than the specified size: displaying a
nucleotide sequence that spans the region, and displaying the
locations of one or more variations in the nucleotide sequence.
6. The method of claim 1, wherein the plurality of variations
includes copy number variations (CNVs), small nucleotide variations
(SNVs), and loss of heterozygosity (LOH).
7. The method of claim 1, wherein the display presents separate
graphs across the region of interest for copy number, allele
specific copy number, and protein encoding sequences.
8. The method of claim 1, wherein a user may select a karyogram
view, a circular view across one or more chromosomes, or a linear
view of a region of interest, any of which indicate where
variations in complex sequence occur.
9. The method of claim 1, wherein step (a) comprises: (i) receiving
a first identifier identifying a user and a second identifier
identifying a test sample; (ii) determining whether the user has
authority to access sequence data for the test sample; and if so,
(iii) retrieving complex sequence for the test sample from a secure
database, the secure database including genomes for a plurality of
different samples.
10. The method of claim 1, wherein step (a) comprises sequencing
DNA contained in the test sample, thereby obtaining complex
sequence for the test sample.
11. The method of claim 1, wherein step (b) comprises iteratively:
(i) sourcing complex sequence for any one of a plurality of a
reference samples in a reference database; (ii) comparing the
complex sequence for the test sample with the complex sequence
sourced in step (i); (iii) selecting the reference sourced in step
(i) as a suitable reference for analyzing the complex sequence for
the test sample if it matches the test sample according to preset
criteria.
12. The method of claim 1, further comprising receiving a selection
of a filter that specifies characteristics of sequence variations
to be displayed; and adapting the display to highlight variations
having the characteristics and/or remove variations not having the
characteristics specified by the filter.
13. The method of claim 1, further comprising predicting one or
more effect(s) of each of at least some of the variations
identified in step (c) on protein expression, protein function,
clinical presentation, and/or disease risk, and providing
information about the effect(s) with the gene sequence map in step
(d).
14. The method of claim 1, further comprising comparing at least
some of the variations identified in step (c) with known variants,
and providing information about the known variants with the gene
sequence map in step (d).
15. The method of claim 14, wherein the information about the known
variants has been obtained at least in part from external
databases.
16. The method of claim 1, further comprising receiving a selection
of one or more of the variations in DNA sequence identified in step
(c), and indicating the one or more variations on the comparative
genetic map displayed in step (d).
17. The method of claim 1, further comprising: receiving a
selection of one or more of the variations in DNA sequence
identified in step (c), determining the position of each of the
selected variations in the human genome, obtaining information
about DNA sequence variations observed for other samples at or
around the position for at least some of the selected variations,
and providing a display comprising the information.
18. The method of claim 1, further comprising: receiving, at the
user interface, an annotation for one or more of the variations in
DNA sequence identified in step (c); and displaying the annotation
on the comparative genetic map displayed in step (d).
19. The method of claim 1, further comprising: receiving from a
second user interface an annotation for one or more of the
variations in DNA sequence identified in step (c); and indicating
the annotation on the comparative genetic map displayed in step
(d).
20. The method of claim 18, further comprising: providing on the
comparative genetic map displayed in step (d) at or near each of
one or more of the variations in DNA sequence identified in step
(c) a hyperlink to a forum whereby a plurality of users having
access to the complex sequence may exchange first information and
reply information regarding the respective variation.
21. A computer product comprising a non-transitory computer
readable medium storing a plurality of instructions that when
executed control a computer system to perform the method of claim
1.
22. A system configured to calculate and provide a display of
variations in complex sequence data of a test sample relative to
complex sequence data for a reference sample according to the
method of claim 1.
23. The system of claim 22, comprising a computer processor, a
database storing the reference DNA sequence data, and a DNA
sequencer.
24. The system of claim 21, wherein the system is configured so
that the user may share information with others about one or more
variations identified in step (d).
25. A server system for sharing variations in complex sequences
between clients, the server system comprising one or more
processors configured to: assemble a gene sequence map that shows a
plurality of variations between a sample complex sequence for a
test sample and a reference complex sequence for a reference
sample; provide the gene sequence map to a plurality of clients;
provide a first user interface to a first client, wherein the user
interface is configured to: receive directions to make the gene
sequence map available to other clients; and receive first
information about one or more of the variations made available to
the other clients; provide a second user interface to one or more
second clients; receive reply information from the one or more
second clients about the one or more variations made available; and
provide, to the first client, the reply information for the one or
more variations.
26. The server system of claim 25, wherein the gene sequence map is
provided to the plurality of clients with links from one or more of
the variations to first information and reply information for the
respective variation.
27. The server system of claim 25, wherein the first information
about the one or more variations provided by the first client user
is made available to other clients with hyperlinks that link to an
indication of the one or more variations on the gene sequence
map.
28. The server system of claim 27, wherein the first information is
displayed in a feed as one or more entries, the feed being
available to a plurality of users, wherein each entry corresponds
to a respective variation.
29. The server system of claim 27, wherein the first information
about the one or more variations is made available to the other
users by way of an information website on the Internet.
30. The server system of claim 28, wherein the server is configured
to compile reply information about one or more of the variations
from other users into an assessment as to whether the variation(s)
are pathogenic or benign.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is a non-provisional of and claims the
priority benefit of U.S. provisional application 61/717,773, filed
Oct 24, 2012. The priority application is hereby incorporated
herein in its entirety for all purposes.
FIELD
[0002] This disclosure relates generally to the field of genome
sequence analysis. More particularly, computer systems and methods
are described that provide a graphical display of sequence
variations and that can be integrated with a network to promote
publication and community feedback.
BACKGROUND
[0003] Methods for whole genome sequencing have been described, for
example in U.S. Pat. No. 7,910,354 (arrays of amplified
polynucleotides), U.S. Pat. No. 8,278,039 (random array DNA
analysis by hybridization); US 2009/0318304 A1 (efficient shotgun
sequencing methods); and US 2011/0033854 A1 (long fragment read
sequencing). Methodology for assembling DNA sequence fragments into
extended genomic sequences includes but is not limited to the
methodology described in U.S. Pat. No. 8,053,191 (iterative nucleic
acid assembly using activation of vector-encoded traits); US
2011/0004413 A1 (calling variations in a sample polynucleotide
sequence with respect to a reference polynucleotide sequence); US
2009/0105961 A1 (methods of nucleic acid identification in
large-scale sequencing); and US 2008/0221832 A1 (methods for
computing positional base probabilities).
[0004] Besides determining a genome, it is beneficial to be able to
visualize the genome in an efficient and intuitive manner.
BRIEF SUMMARY
[0005] This disclosure provides a computer system and method for
determining, displaying, and discussing variations that occur in
genetic sequences between different individuals.
[0006] One aspect of the invention is a method for displaying
variations between genomes between different individuals of the
same species, and differences between other complex sequences.
Sequence data is received for a test sample and a reference sample,
the samples are aligned using a computer system, and variations in
sequence are identified. The variations can then be displayed on a
comparative gene sequence map showing where the variations are
located. The user can be provided with an interface for controlling
display of the comparative gene sequence map. The user can select
from the interface a selection of a region within the genetic
sequence map (sometimes less than half the entire comparative gene
sequence map), whereupon the display is reconfigured such that the
region is magnified, optionally in proportion to the size of the
region: for example at least two times the original display,
presented across at least about one half of the width of the
display.
[0007] A display system according to the invention may be
configured to display sequence and variant data in several formats.
The formats may include a karyogram view, a cytogenomic view, a
chromosome linkage view, a chromosome view, a linear map, or a
sequence view, typically with any combination of several or all of
these formats available and user selectable.
[0008] The gene map can provide resolution down to the level of
single bases. Thus, the method comprises determining whether the
region is smaller than a specified size, and if so, displaying a
nucleotide sequence that spans the region, and (should at least one
of the determined variations occur within the selected span at the
specified size), displaying the locations of the variations with
respect to the nucleotide sequence. Gene variations for display
according to the method include but are not limited to copy number
variations (CNVs), small nucleotide variations (SNVs), loss of
heterozygosity (LOH), and other variation types listed elsewhere in
this disclosure.
[0009] Richness of the display can be adapted to the liking of the
provider and/or the user. For example, the display can present
separate graphs across the region of interest for copy number,
allele specific copy number, and protein encoding sequences. The
user may be given the capacity to select a karyogram view, a
circular view across one or more chromosomes, or a linear view of a
region of interest, any of which indicate where variations in
complex sequence occur.
[0010] To provide secure access to complex sequences of test
samples, the method may comprise receiving a first identifier
identifying a user and a second identifier identifying a test
sample, determining whether the user has authority to access
sequence data for the test sample, and if so, retrieving complex
sequence for the test sample from a secure database, the secure
database including genomes for a plurality of different samples.
The method may also comprise doing the actual sequencing of DNA
contained in the test sample, using any suitable method, such as
those listed elsewhere in the disclosure. The reference sequence
can optionally be selected from a database of such sequences, for
example, by sourcing complex sequence for any one of a plurality of
a reference samples in a reference database; comparing the complex
sequence for the test sample with the reference sequence, and
selecting a reference sequence if it matches the test sample
according to preset criteria.
[0011] Filtering of the data can be done by receiving a selection
of a filter that specifies characteristics of sequence variations
to be displayed; and adapting the display to highlight variations
having the characteristics and/or remove variations not having the
characteristics specified by the filter. The method may comprise
predicting one or more effects of sequence variations on protein
expression, protein function, clinical presentation, or disease
risk, and providing information about the effects with the gene
sequence map. The method may also compare at least some of the
variations with known variants, and providing information about the
known variants with the gene sequence map, optionally from external
databases. Certain variants can be highlighted or pinned by the
user by receiving a selection of one or more of the variations in
DNA sequence, and indicating the one or more variations on the
comparative genetic map. The method may comprise receiving a
selection of one or more of the variations in
[0012] DNA sequence, determining the position of each of the
selected variations in the human genome, obtaining information
about DNA sequence variations observed for other samples at or
around the position for at least some of the selected variations,
and providing a display comprising the information.
[0013] Another aspect of the invention is a computer product
comprising a non-transitory computer readable medium storing a
plurality of instructions that when executed control a computer
system to perform any aspect of the aforedescribed method.
[0014] Another aspect of the invention is a system configured to
calculate and provide a display of variations in complex sequence
data of a test sample relative to complex sequence data for a
reference sample according to methods described herein. The system
may comprise a computer processor, a database storing the reference
DNA sequence data, and a DNA sequencer. The system may be
configured so that the user may share information with others about
one or more variations.
[0015] Another aspect of the invention is a server system for
sharing variations in complex sequences between clients. The system
comprises one or more processors configured to assemble a display
such as a gene sequence map that shows a plurality of variations
between a sample complex sequence for a test sample and a reference
complex sequence for a reference sample. The processors provide the
gene sequence map to a plurality of clients; provide a first user
interface to a first client, wherein the user interface is
configured to receive directions to make the gene sequence map
available to other clients, and receive first information about one
or more of the variations made available to the other client. The
system is configured to provide a second user interface to one or
more second clients; receive reply information from the one or more
second clients about the one or more variations made available; and
provide, to the first client, the reply information for the one or
more variations.
[0016] The system may provide the gene sequence map to the
plurality of clients with links from one or more of the variations
to first information and reply information for the respective
variation. The first information about the one or more variations
provided by the first client user can be made available to other
clients with hyperlinks that link to an indication of the one or
more variations on the gene sequence map. The first information can
be displayed in a feed as one or more entries, the feed being
available to a plurality of users, wherein each entry corresponds
to a respective variation. The first information about the one or
more variations can be made available to the other users by way of
an information website on the Internet. The server can be
configured to compile reply information about one or more of the
variations from other users into an assessment as to whether the
variations are pathogenic or benign.
[0017] Other aspects of the invention will be evident from the
description that follows.
Definitions
[0018] The terms "complex sequence", "sequence compilation" or
"target sequence" as used in this disclosure refer to a set of
nucleotide sequences for an individual or combination of
individuals of any species. The sequence data may be obtained by
sequencing a biological sample from a particular individual, or it
may be constructed as artificial or consensus sequence data for a
human individual or population. The complex sequence may be
complete or partial genome sequence data, it may be cDNA sequence
data from an expression library, it may be a sequence collection
such as an exome or a biome, or it may be another DNA compilation
of interest to the user. Unless explicitly stated otherwise, a
suitable individual from which the complex sequence has been
obtained may be selected from a human, other mammals, other
vertebrates, other eukaryotes, prokaryotes, or combinations
thereof, including but not limited to a pool of human or other
biological samples from a particular source or from a particular
human subclassification, single celled organisms from a particular
environment, or a compilation of consensus or hypothetical
sequences of an organism or component thereof.
[0019] A "reference sample" as used in this disclosure is any
sample to which the user may compare a test sample, as explained
below. The reference sample can have a reference complex sequence,
e.g., a reference genome. A sequence of a nucleic acid molecule
from the test sample (thus corresponding to the genome of the test
sample) can be aligned to the reference sequence.
[0020] A "comparative gene sequence map" is a graphical depiction
in which variations between two or more complex sequences are
mapped between genetic position indicators (such as base count or
known markers) along a DNA structure such as a chromosome. Unless
otherwise specified, gene sequence maps provided for display are
"expandable", in the sense that the user may request depiction of
an overview (such as an entire chromosome), or a segment thereof in
increasing detail, ultimately resolving into the actual nucleotides
in the sequence in the selected segment.
[0021] Nucleotide sequence "variations" include any difference
between a complex sequence (e.g., a genome) of one sample and the
complex sequence of another sample, as determined from sequence
reads of the respective samples, which is explained and exemplified
below.
[0022] A "display" is a visual depiction shown on an I/O device
such as a monitor or touchpad, or within a frame or window depicted
by the device. A computer system may "provide a display" of an
item, for example, by causing the item to present on a suitable
output device operatively connected to the system, or by sending to
an external device (such as a client) a set of data that will
enable the device to present the item locally.
BRIEF DESCRIPTION OF THE DRAWINGS
[0023] FIG. 1A is a block diagram with an exemplary configuration
for a gene explorer system according to this invention. FIG. 1B is
a flow chart showing a process by which a user may have the system
analyze and display sequence data.
[0024] FIG. 2A is a screenshot showing a display of genome sequence
data from a particular patient in a karyogram view. FIG. 2B shows a
cytogenomic view of the entire genome, compared with a reference
sequence.
[0025] FIG. 3A is a display showing an overview of the entire
genome sequence of the patient sample in the form of a
Circos.RTM.-style plot, showing analysis of sequence variations and
apparent interchromosomal junctions. FIG. 3B is a more detailed
view mapping nucleotide variations determined by comparing a test
sequence with a reference sequence.
[0026] FIG. 4A is a display of Chromosome 1 in linear view, which
can be expanded by moving the markers 402 to a narrower region for
more detailed viewing. FIG. 4B shows a screenshot in which the user
calls up a keyboard to input the name of a particular gene of
interest for an exploded view. FIG. 4C shows a subregion of
Chromosome 17, indicating the positions of genes TP53 and
WRAP53,and single nucleotide variations (SNV). FIG. 4D shows a
further expanded view of a particular gene on Chromosome 14. At
this level of expansion, individual nucleotides of the genomic
sequence are shown. The test sequence and reference sequence are
the same, except where there is a variation: in this case, an SNV
437.
[0027] FIG. 5A shows a menu to filter the analysis towards specific
genes or specific chromosomal locations of interest or potential
importance. FIG. 5B shows a menu for assembling a gene panel. FIG.
5C shows a menu where the user selects chromosomal regions by
function of the nucleotide sequence (for example, encoding regions,
non-encoding regions within genes, and sequence between genes).
FIG. 5D shows a menu that allows the user to select variations
depending on the effect on the encoded protein.
[0028] FIG. 6 shows a display with a summary of the analyzed data
for selected genes characterized the effect of the nucleotide
variations on the encoded protein.
[0029] FIG. 7 shows a display in which a particular sequence
variation has been evaluated by a community of users.
[0030] FIG. 8 shows a social networking portal that has been
configured for promoting a dialog between a user that has analyzed
a sequence for variations according to this invention, and other
users who may have an interest in the variations observed or the
genes in which they have occurred.
[0031] FIG. 9 is a flow chart showing an exemplary process of this
invention by which a user may submit a sequence for processing and
obtain a display with the resulting analysis.
[0032] FIG. 10 shows the subsystem architecture of a computer by
which a gene explorer system according to this invention may be
implemented.
DETAILED DESCRIPTION
[0033] This disclosure provides a technology for users to gain
first-hand knowledge and experience with interpreting whole
genomes. The technology graphically depicts variations in genome
sequences in an expandable display, and provides a platform whereby
the user may find and research the biological significance of such
variants. The technology also provides a unique collaborative
environment designed to capture and improve the collective
knowledge of the participating community.
[0034] By way of illustration, each user may analyze one or more
complex sequences in the following fashion. After presenting their
credentials, the user calls from a client for a complex sequence
obtained for a test sample and stored on a secure database
accessible by the sever. The system matches the test sequence to a
selected reference sequence. The test and reference sequences are
aligned, and sequence variations are determined. The variations are
mapped on the sequence in a graphical display which is provided to
the user by way of the client. The user may explore regions of
interest on the map by directing the system to move about and
expand the display. The user can drill down to review gene
variations at the nucleotide level. The user can also request that
the data be filtered for particular genes or particular types of
variations.
[0035] The computer system also provides a platform by which the
user may authorize other users to have access to the test sample
sequence, and the genetic variations determined therefrom. Both the
primary user and other users may annotate the variations and
exchange comments over a social network that is integrated into the
system.
I. Overview
[0036] Whole human genome sequencing holds great promise as a
diagnostic and predictive aid in the management of human health.
However, interpreting whole genome data to obtain clinically
instructive information poses substantial challenges. Before
development of the technology described here, expertise in
analyzing and interpreting genome sequence data has been
distributed across a variety of genomic databases, making it
difficult to assemble different sorts of information and resolve
open questions by consensus. The technology described here
addresses these challenges by consolidating information from
multiple sources, and providing a platform for more users to learn
and share knowledge about interpreting genomic variants.
[0037] The technology described here provides a number of important
advantages over what was previously available. First, the sequence
analysis system described here can be integrated with a service for
determining genomic sequence of a biological sample. The new
sequence data can be consolidated with information from multiple
sources, including prediction algorithms and cross-reference
annotations from external genomic databases. Second, the technology
helps users interpret genomes by providing users with important and
instructive information through an easy-to-use interface.
[0038] Third, the technology allows experts to collaborate across
the platform. In the tradition of modern electronic networks where
people connect and share ideas in business and their personal
lives, the technology described here has a collaborative function
specifically focused on the interpretation of genome sequence
variants This creates a vibrant knowledge base in genomics, to the
benefit of clinician researchers everywhere and the patients under
their care.
[0039] FIG. 1A is a block diagram showing an illustrative
configuration for a gene explorer system 100 according to this
invention A first user accesses the system through a first client
computer or terminal 103 operatively connected to a local or
network accessible server 104. The client 103 has access to and
provides sequence data from a database 102. Typically, the database
102 is secured to protect confidentiality or control access of
sequence data obtained from a sequencing service 101, obtained for
example by sequencing one or more biological test samples. The
secure sample sequence database 102 may be connected independently
to the client computer 103, or it may be a partition of a shared
database connected with the server 104 to which the user controls
access. The server processes data from the database 102 so as to
create a display for presentation on the client computer 103. The
processing typically includes comparing the test sequence with one
or more reference sequences, such as a consensus sequence in a
region of interest with respect to a particular disease or
condition. The reference sequences used in the analysis may be
stored on a reference sequence database 107 compiled by the server,
and/or obtained from an external source 108.
[0040] Each variation identified between a test sample and a
reference sample may be analyzed by the server 104 with a plurality
of filters or algorithms (for example, to determine if it causes a
change in the encoded protein), and annotated accordingly. Each
variation may be compared with previously known variations, and
annotated according to the clinical or academic significance of the
previously known variant. The annotations (or a selection thereof)
are each provided at or near the respective sequence variation
displayed on the client computer 103. In some configurations of the
invention, the user has the option of making the test sequence
available to one or more other users having their own client 105
connected to the server 104. Each of the other users may or may not
bring their own sequence of interest from a client-specific
database 106 for purpose of comparison or analysis. Information
from the other clients 105 may be used by the server 104 to
annotate the display presented on the first client 103. The server
104 may also provide a forum whereby the first user and the other
users may exchange written commentary with regards to variations
observed in the test sample.
[0041] FIG. 1B is a flow chart showing a process by which a user
may submit a sequence for processing and obtain a display according
to this invention. Features of this process are described below in
Section III.
[0042] The methods and systems described here can be used for a
variety of research purposes and techniques. One such application
is to assist a user in evaluating genetic variations that may be
present in the genome, diseased tissue, or expression library of a
particular individual. Such information may be instructive in the
clinical management of the individual's underlying condition.
[0043] Thus, the user may obtain and upload a complex sequence from
test sample into the system before analysis. Another application of
the technology is to integrate the analytical tools with a DNA
sequencing service, whereupon sequence information obtained from a
particular individual becomes the subject of the analysis.
[0044] Sequencing methodology suitable for use with this invention
includes but is not limited to the methodology described in U.S.
Pat. No. 7,910,354; U.S. Pat. No. 8,278,039; US 2009/0318304 A1;
and US 2011/0033854 A1. Generally, complex sequence is fully
assembled before being used to identify sequence variations as
described here. Methodology suitable for assembling DNA sequence
fragments into extended genomic sequences includes but is not
limited to what is described in U.S. Pat. No. 8,053,191; US
2011/0004413 A1; US 2009/0105961 A1; and US 2008/0221832 A1, which
are here incorporated herein by reference in their entirety for all
purposes.
[0045] Complex sequence from a test sample can be loaded from the
sequencing apparatus into the analytic system of this technology
automatically, by the system operator, or by the end user. The
system can be configured with a security system so that each user
may control access to complex sequences from each sample. The user
may be provided with the sequence data in electronic form, and then
the source data is expunged from the server. More conveniently, the
server may provide a repository of data, so the user may access the
data on the server by cloud computing, and thereafter apply the
analytic tools of this invention to the data from each test sample
at the user's convenience.
[0046] To maintain access control, for example, the server can be
programmed to receive a first identifier identifying a user and a
second identifier identifying a test sample. It then compares the
identifiers with stored information to determine whether the user
has authority to access sequence data for the test sample. If the
requesting user has such authority, the server then retrieves
complex sequence for the test sample from a secure database, and
permits the user to view, analyze, and optionally download the
data. The user may also have authority to change permissions, such
that access to the complex sequence data and/or the subsequent
analysis is opened up to other specified users or class of users,
other members of having access to data on the server, or the public
at large.
II. Identification and Display of Sequence Variations
[0047] In order to determine variations in the sequence, the system
can compare the complex sequence from the test sample with complex
sequence from a reference standard. After receiving complex
sequence for a test sample, the system aligns the complex sequence
for the test sample with complex sequence for one or more reference
samples, so that substantially identical portions of sequence are
compared. Differences between the test sequence and the reference
standard are identified by the system as variations, which may then
be filtered as described below as an aid to evaluating their
biological and clinical significance.
[0048] The reference standard used for this analysis may be
consistent for all test samples during a period of operation.
Alternatively the reference standard may be provided or identified
by the user for the analysis of a particular test sample. This may
be useful, for example, in the analysis of tissue from a tumor,
where normal or unaffected tissue from the same person may serve as
the closest reference.
[0049] In some implementations, the server will have access to a
plurality of potential reference standards, and the server will
identify one or more references to be used for each test sample
analyzed by the system. In this implementation, the system will
source complex sequence for any one of a plurality of standard
samples in a reference database. It will compare the complex
sequence for the test sample with the complex sequence called from
the reference base. It can then select a standard from the database
if the standard sample matches the test sample according to preset
criteria.
[0050] A. Identification of Sequence Variations
[0051] Methods and systems of this invention can be used to
identify variations in complex sequence. These are differences
between the complex sequence of one or more test samples and one or
more reference samples.
[0052] Nucleotide sequence variations include any difference
between a sequence determined for one sample and the genome of
another sample, as determined from sequence reads of the respective
samples. Apparent variations may arise from errors in sequencing
and/or sequence assembly. Where a variant is known or suspected of
being a sequencing error, it may be suppressed or de-emphasized in
the map displayed to the user. True sequence variations between a
test sample and a(those reproducible upon resequencing or otherwise
suspected or confirmed) include but are not limited to the
following: [0053] Small nucleotide variations (SNVs) and single
nucleotide polymorphisms (SNPs). SNVs include the change, addition
or deletion of one or more nucleotides in or around the region of
variation. SNPs are a subset of SNVs wherein the variation of a
single nucleotide occurs with known regularity between different
individuals or different types of samples of the same species. SNPs
include inheritable alleles and mutations in diseased tissue that
occur with known regularity. [0054] Copy number variations (CNVs),
where a particular gene or other functional element of a complex
sequence is replicated or reduced in replicates compared with a
reference sample, such as may occur through gene duplication and/or
translocation. [0055] Loss of heterozygosity (LOH), chromosome
trisomy and gene rearrangement (transposition). Embodiments of the
invention can cause any one, some or all of such types of
variations to be displayed in any combination.
[0056] The reference sample may be any sample to which the user may
compare a test sample. It may be a sample of another individual,
but is more typically a consensus or hypothetical sequence, such as
may be created by amalgamating the corresponding sequences of a
plurality of individuals, and/or editing to exclude or include one
or more nucleotide variations for the purpose of the comparison.
The comparison can be done by aligning the test sequence with the
reference sequence so as to minimize the number of variations. For
example, the test sequence can be aligned to a location that
results in the fewest mismatches or variations from the reference
sequence.
[0057] The reference sample may be selected from a database of
reference samples to match the sample being analyzed. A reference
sample is chosen to match a test sample if it is sufficiently
similar to the test sample in terms of one or more preset criteria.
By way of illustration, determining a match may include any one or
more of the following criteria in any combination: degree of
identity, present or absence of particular sequence variations of
interest, and source information such as racial background, family
relatedness, and clinical condition.
[0058] B. Display of Sequence Variations
[0059] The technology described here provides a platform by which
users may visualize, filter and analyze variations in genomic
sequence data in one consolidated tool. For example, small
nucleotide variations (SNVs), copy number variations (CNVs) and
loss of heterozygosity (LOH) events can all be viewed on the same
display. A built in genome browser is provided to visualize and
surf genomes with genome wide views down to base pair level views,
so that the user may explore sequence variations that are relevant
to their particular clinical situation.
[0060] As an introduction to results of the analysis and to orient
the user, the system can provide the user with the choice of
various views of the genome or expression library as a whole.
[0061] By way of illustration, FIG. 2A is a screenshot 200 showing
a display of genome sequence data from a particular patient. On the
right side is a karyogram view 210, with copy number variations
shown to the right of each chromosome 211, depicted according to
its banding pattern 212. Copy number variations are shown to the
right of each chromosome: Copy number gain, shown as downward
pointing chevrons 213, copy number loss, shown as upwards pointing
chevrons 214, and loss of heterogeneity, shown as horizontal hash
marks 215.
[0062] The display on the left side 220 provides a list of
particular variations in a panel of genes of interest 221 which a
user has selected for analysis, with a summary assessment
underneath 222 as to whether it is pathogenic or benign. The gene
panel is selected as described below, in reference to FIGS. 5A and
5B. Each variation is named 221 according to its mapped position in
the reference sequence, and the change made. Thus, c.29085809A>G
is a variant detected on Chromosome 22 of the reference sequence
mapping to position 29,085,809, where the nucleotide A (alanine) in
the reference sequence is changed to G (glutamine) in the test
sequence. Prediction of whether a particular variation is likely to
be pathogenic 222 is made on the basis of the expected change to an
encoded protein sequence, experience of such a change in other
instances, or other types of predictive analysis in any
combination. The display also comprises an icon 223 by which the
user may call up further details, such as the test sequence
accession number, the reference sequence accession number or
designation, the position, the gene or sequence common name or
designation, its symbol, further information on the type of
sequence variation or SNV, and the zygosity (homozygous or
half).
[0063] FIG. 2B is another non-limiting illustration showing a
screenshot 230 comprising a cytogenomic view. On the right side is
a cytogenomic view 240 of the entire genome. Here, the data for
each patient is compared with a sequence standard drawn from CGI's
database. The top track 241 is copy number relative coverage data,
which is the number of sequence reads of each region as a ratio of
the coverage of the sample. The next track 242 is allele specific
copy number which may show loss of heterozygosity (LOH) in a
particular region due to a gene deletion or rearrangement event
(for example, in cancer cells). The third track 243 shows a count
of small nucleotide variations (which includes SNPs, inserts,
deletions, and small substitutions) that occur in the gene panel
selected by the user. The gene track 244 shows the concentration of
genes across the genome. As the user zooms in to a particular
region of interest, they will see the actual gene names as well as
the exons and introns. On the left (as in FIG. 2A), there is a
display 250 of known variations in certain genes of interest 251
that have been selected as a gene panel for assessing a particular
condition, and a graphical assessment 252 as to whether the
variations are predicted to be pathogenic or benign.
[0064] FIG. 3A is an overview display 300 of the entire genome
sequence of the patient sample in the form of a circular or
Circos.RTM.-style plot (M. Krzywinski et al., Genome Res.
19:1639-1645, 2009). This display shows the frequency of structural
variations at locations across the chromosome map. Curved lines in
the middle 301 show apparent interchromosomal junctions. FIG. 3B
provides a detail 310 for Chromosome 4. From the outside in, the
layers show a chromosome map 311, the called ploidy 312, the lesser
allele fraction 313 (the fraction of the sample containing the
allele that is present in .ltoreq.50% of the sample), heterozygous
SNP density 314 (density of occurrence of SNPs as a single
haplotype), and homozygous SNP density 315 (density of occurrence
of homozygous SNPs).
III. Expandable Display
[0065] One aspect of the invention is a display system for
surveying and analyzing complex sequence data. Referring back to
FIG. 1B, the display system can receive sequence data 151, for
example, new data inputted or transmitted to the system from a
sequencing service that has determined the sequence of one or more
biological samples of interest. One or more than two test sequences
are aligned with one or more reference sequences 152 that are
either standard to the analysis, or selected by the system or by
the user as an appropriate match for the test sample. Such
references may be stored within a database of reference sequences
that have been stored or compiled by the system and/or retrieved
from one or more external sources. The system may optionally update
one or more reference sequences in the database on an ongoing basis
as a learning function in relation to test sequences that are
processed by the system
[0066] Once the samples are aligned, the system then processes the
data to determine where there are variations 153 between the test
sequence(s) and the reference sequence(s). Variations include but
are not limited to small nucleotide variations (SNVs), copy number
variations (CNVs), loss of heterozygosity (LOH), and
translocations. The variations that are identified by the system
can optionally be filtered 154 to facilitate interpretation of the
analysis. For example, one or more filters cam be implemented to
exclude variations identified or suspected to be sequence read
errors or assembly errors. As described elsewhere in this
disclosure, other filters that can be implemented by choice of the
use include but are not limited to filters that focus the display
on particular genes or gene regions, filters that select variations
depending on their effect on the function of the sequence (for
example, the effect on an encoded amino acid sequence), and filters
that select variations depending on previously known variants of
particular interest or clinical significance. Such filters can be
used singly or in any effective combination, and can be switched on
and off by choice of the user during her review of the displayed
data.
[0067] A display is then provided by the system 155 to a user
client that show the filtered variations at their location on a map
of the complex sequence: all the way from an overview map of the
entire sequence (such as a karyogram view for a genomic sequence),
in various degrees of detail down to the actual nucleotide
sequence. The display is user controllable in the sense that
(depending on how the system is configured), the user may request
one or more of the following in any combination: a change in
expansion or detail, the imposition or removal of filters,
highlighting or annotating a particular variant identified by the
system or a gene region comprising one or more variations of
interest, and permitting other system users to provide their own
annotations directly on the display or in a linked forum. In the
configuration shown here, the user interface receives a selection
of a region within a larger sequence map 156, whereupon the system
reconfigures the display to magnify or focus on the region selected
by the user. The user interface can also receive a request to
decrease magnification or expand the scope of the region being
analyzed, whereupon the system reconfigures the display to compress
or scale down the previously viewed region into a smaller space so
as to provide space for neighboring regions of the complex sequence
to appear in the display.
[0068] FIGS. 4A, 4B, and 4C illustrate the capacity of the display
to drill down to subregions of interest to the user to inspect
details of each sequence variation. The benefit of the expandable
display is that it reconfigures in real time in response to user
input so as to provide an expanded view of a particular region of
interest. Thus, a genomic overview (such as FIG. 2A, FIG. 2B or
FIG. 3A) can be expanded to detail a particular chromosome (FIG.
4A), shown here in a horizontally linear depiction. This in turn
can be expanded to a particular subregion or gene cluster (FIG.
4C), and then to the area around and about one or more sequence
variations of interest (FIG. 4D). The system maintains sequence
data as the basis of the graphical display. Thus, if a sufficiently
narrow region of interest is selected, the display will present
nucleotide sequence on a nucleotide-by-nucleotide basis spanning
the narrow region of interest. In this display, a small nucleotide
variation (SNV) 437 can be identified as a particular nucleotide
deletion, addition, or substitution in relation to the reference
sequence.
[0069] Navigation between the views may be implemented in any way
that is compatible with the underlying code and hardware. For
example, the user may be provided with a numerical input or sliding
scale to specify the degree of zoom from the original depiction. If
the I/O device has a touch screen, then the view can be expanded,
contracted, or moved up or downstream by touch, pinch, zoom, or tap
at appropriate locations in the previous view. The interface may
also be provided with links or menu choice whereby the user may
specify a particular region, gene, variation, or other chromosomal
feature, and be taken to a view whereby the selected feature
occupies at least 10%, 20%, 40%, a majority, or substantially all
of the width of the display. The system can also be configured so
that the user has the option of pulling up a keyboard (FIG. 4B)
through which to input the name of a particular gene or region upon
which the display should be focused.
IV. Filtering and Data Interpretation
[0070] Genomes from different individuals who are not related
inevitably comprise a range of sequence variations that provide a
genetic basis for their individuality, but which do not necessarily
portend a particular phenotype or disease susceptibility. Depending
on the quality of the sequencing, there may also be sequencing
errors that create noise potentially obscuring important data. To
aid the clinician researcher in focusing on the variations more
likely to be clinically significant, the system can provide a
filtering capability to narrow the range of exploration.
[0071] FIGS. 5A, 5B, 5C, and 5D provide some non-limiting examples
of the types of filters that can be implemented. In FIG. 5A, the
user may focus the analysis towards certain genes 501 or specific
chromosomal locations 502.
[0072] In FIG. 5B, the user positively selects particular genes or
gene categories of particular interest, from amongst a broad or
comprehensive list of encoding regions, thereby forming a "gene
panel". In this illustration, a panel of 15 genes known to be
relevant for certain diseases or clinical conditions has been
selected. The user has the option of selecting lists of between 1
and 10,000 genes, between 3 to 10,000 genes, between 5 to 1,000
genes, or between 10 and 100 genes. Instead of or in addition to
selecting genes associated with a particular disease or condition,
genes may be selected for the panel according to a selected
functionality: for example, genes that encode cell surface
receptors, or cell surface receptors specific for a certain ligand.
The system can be configured to assist the user in selecting
particular genes, or allow the user to select the entire category.
In a similar fashion, the filter may permit the user to focus on
non-encoding regions thought to be involved in another aspect of
gene regulation, DNA processing, or biological phenotype. As
another option, rather than selecting the genes one by one, the
user or the system may utilize a predefined gene panel associated
with a particular disease or condition of interest.
[0073] FIG. 5C is another type of filter provided by the system
that allows the user to focus on particular chromosomal regions
according to their function--either for genes positively selected
as part of a panel, or across the entire genome. Thus, the user can
select encoding regions (CDS), exons, untranslated regions (UTR) at
the 5' end, the 3' end, or both, transcription regulators such as
promoter or enhancer regions, and so on. FIG. 5D is a filter that
allows the user to focus on particular types of genetic variations
in terms of their predicted effects on the encoded protein. For
example, small variations within an encoding sequence can be
categorized as follows: [0074] a gene alteration that results in
selection of a codon synonym (hence no change on the encoded amino
acid), [0075] in frame effects--e.g., alterations, deletions, or
insertions that cause point variations but otherwise leave the
encoded protein sequence intact, or [0076] more profound changes
likely to compromise protein function, especially if they occur
early in the coding sequence (for example, frame shift mutations or
insertion of a stop codon).
[0077] By way of further illustration, small nucleotide variations
(SNV) can be filtered according to predicted pathogenicity, gene
region, impact on encoded amino acid sequence, SNV type (e.g.,
single nucleotide polymorphisms, deletions, insertions,
multi-nucleotide substitutions), zygosity (homozygous or
heterozygous), whether it is found in a particular database such as
dbSNP, or to exclude commonly found variants in healthy
populations. Copy number variations (CNV) can be filtered according
to predicted pathogenicity, gene region, number of copies, repeat
category overlap, or whether it is found in a particular database
such as DGV. Any quality filters, such as ploidy quality score, and
CNV type quality score. Different types of variations can be
screened for quality or confidence of the sequencing and/or the
sequence assembly so that sequencing errors do not confound the
analysis. Such filters may be selected by the user for use in any
effective combination, optionally in combination with a filter of
the user's own devising that has been constructed on or loaded onto
the system.
[0078] The computer system and methods of the invention can be
implemented so as to provide annotations and further information to
assist in drawing conclusions from the presence of genetic
variations that are presented after filtering. For example, the
server may provide algorithms to predict protein expression,
protein function, clinical presentation, and/or disease risk in any
combination. Protein expression can be predicted on the basis of
the effect on the encoded amino acid sequence. Protein function can
be predicted on the basis of predicted three-dimensional structure,
and effects on catalytic and binding sites. Clinical presentation
and disease risk can be predicted based on cumulative history of
other known variations at or around the same location and their
known effects. Such predictive algorithms can be programmed into
the system, or sourced from external servers, for example, over the
Internet.
[0079] The computer system and methods of the invention can also be
implemented so as to provide comparisons with gene variations
previously reported that either occur in the same genes or that
have similar biological and/or clinical effects. The system can
source such information from within its own records of similar
analyses done on other complex sequences in the same customer and
reference databases. The system can also source such information
from other systems and databases that are external to the system,
for example, over the Internet. Once such other known variations
are identified, the system can indicate the position of each
variation on the same genetic map and/or provide a nearby link to
information known about each other known variant. The annotation
function can be provided as both a passive and an active tool.
[0080] Thus, upon receiving information on user preferences, the
system may as a matter of course annotate the genetic map with
information about nearby or related genetic variants using internal
and/or externally obtained information, in accordance with the user
preferences. Alternatively or in addition, the system may be set up
with a query function. Thus, the user may select on one or more
genetic variations in a test sequence, or one or more gene regions
in the test or reference sequence. The system may then be prompted
to run its own algorithms and/or seek external information in
accordance with the request from the user.
[0081] The technology described here allows the user to take
advantage of rich variant annotations. This can include (1)
clinically meaningful variant names that follow HGVS standards; (2)
gene annotations by transcript including functional impact and
scores from prediction algorithms such as Polyphen and SIFT; (3)
cross-reference annotations from several genomic databases such as
dbSNP and DGV and (4) allele frequency information across several
data sets such as the 1000 Genomes. The user can thereby be
provided with consolidated information with smart links to
additional resources. Information includes variant attributes,
variant and gene annotations, technical performance summaries and
context sensitive links including links to NCBI, PubMed and popular
public genome browsers. Consolidated views help shorten the variant
analysis and research process.
V. Social Networking
[0082] Another feature that can be implemented in the computer
systems and methods of this disclosure is a platform for
interacting with other users so as to improve the knowledge base,
both with respect to a particular test sequence, and generally in
the study of variations of a particular type or class.
[0083] This can be implemented using a server that provides
information for multiple clients. In reference to FIG. 1A, each of
the other clients 105 is provided with a display of a gene sequence
map assembled by the server 104 for presentation to users, the gene
sequence map showing variations in complex sequence for a test
sample compared with complex sequence for one or more reference
samples (such as in FIGS. 2A and 2B). An interface is provided
whereby a first client user 103 can direct the server to make the
gene sequence map, particular sequence data, or particular genetic
variants available to other client users 105. Optionally, the first
user annotates or selects particular genetic variations or gene
regions of interest, and/or poses particular questions, for
example, with respect to possible biological or clinical effect of
one or more identified genetic variations, or generally seeking
other experiences regarding the genetic variations or gene regions
of a similar nature to those they have selected or researched. The
information is annotated or linked to the display at or near the
locus to which the variation has been mapped.
[0084] Having been granted access to the map or information from a
particular test sample or particular genetic variants, other users
on the system 105 (either members selected by the first user,
subscribing members generally, or a member of the public who has
access to the system over the Internet) can reflect on the
nucleotide variations or the questions posed by the first user.
Should they wish to contribute information in reply, the system
provides a portal by which the reply information is transmitted to
the first client user privately, or to the community of users
selected by the first client user for general discussion of the
topic. The portal can be implemented so as to permit the replying
user to add a location marker and/or an annotation to the gene
sequence map, or to append information to an annotation made by the
first user. The portal can also be implemented so as to permit a
discussion to occur through an exchange posting linked to the map
or previous comments, or provided as a separate display whereby the
first client user can receive reply information about one or more
of the variations that the server obtains from one or more of the
other client users.
[0085] FIG. 8 is a screenshot of a sequence analysis system
according to this invention that has been configured for promoting
such a dialog. In this configuration, there is a window or portal
804 where the first user posts an informational overview about the
sequence being displayed, such as the presence and import of
variations detected therein. In another window or portal 803, other
users authorized to have access post reply information, which can
evoke further commentary from the first user and others. The first
user may optionally be authorized by the system to control access
and/or moderate and edit the discussion.
[0086] In more general terms, the system may provide forums of
various types by which users may interact about particular genetic
variations, particular gene regions, particular biological effects,
or particular clinical issues. Such forums may be in the style of a
bulletin board or blog, optionally with links to gene maps, complex
sequence data, or other discussions in any combination, optionally
with various levels of controlled access depending on privacy
issues or proprietary aspects of gene sequence data and/or expert
interpretation thereof. Such forums may also be in the style of a
news feed with links to gene maps, complex sequence data, or other
discussions in any combination, optionally with various levels of
controlled access. News feeds will typically be provided by
subscription, whereby certain users or class of users may be given
access, and may opt in or opt out to receiving the selected feeds
on a periodic or as-needed basis.
[0087] Such forums may also be in the style of a passive knowledge
base or wiki, where the system provides a repository of information
available for research into particular topics or genetic variants.
The knowledge base may have links to gene maps, complex sequence
data, or other discussions in any combination, optionally with
various levels of controlled access, and optionally also provides a
means by which the initiating user of each entry or other users may
have authority to update, depending on their level of access.
[0088] Thus, the social networking aspect of the invention can be
implemented so that as users filter and search for variants, they
can mark variants of interest by highlighting or annotating them to
get back to them easily. Community assessments allow users to
contribute to and leverage community knowledge about the
pathogenicity of variants. As users use the technology to create
variant assessments, they can opt to share them with the community.
Users can then comment on each other's assessments. The next time
that variant is seen in the context of a case, users see a summary
of all shared assessment as well as the detail of each assessment
including who made the assessment, when and all the supporting
notes, evidence and comments. Newsfeeds provide a way for users to
stay on top of the latest activities and collaborate with other
users. Colleagues can comment and collaborate with each other
directly through the newsfeed to exchange information, solve cases
and assess the pathogenicity of variants.
VI. Illustrative Example
[0089] The figures accompanying this description show a
non-limiting illustration of certain aspects of the invention. The
system for analyzing genetic variants shown in this illustration
has been developed by Complete Genomics Inc. for use in commerce
under the service mark Voyager.TM. or Voyager Genome
Explorer.TM..
[0090] Voyager provides an application platform for use on a
computer station, personal data assistant, touch pad, or other
client configured for interaction with a server that provides
sequence information and display. The platform may be implemented
on standard operating systems such as iPad.TM., iOS.TM.,
Android.TM., and Windows.TM.. The user opens up the application,
logs in, and gets access to cases from the server to which the
logged in user is permitted access. Voyager is integrated
seamlessly with Complete Genomics' gene sequencing service. Thus,
the credentialed user will have access to sequence data from
samples they provided, sequence data that has been made generally
available to Voyager subscribers and/or the general public.
[0091] A user having a complex sequence they want analyzed logs
into the system once the sequence is available from the gene
sequencing service. The sequence is labeled with information about
the test sample: for example, nature of the sample (such as
peripheral blood), the sampling date, and an identifier (ID) for
the particular sample. The sequence is given its own Internal
Accession Number that is unique to the sample and the sequencing
run. Thus, samples from different patients, multiple samples from
the same patient on different days, and different sequence runs for
the same sequence are identified separately.
[0092] Upon launching or enquiring about complex sequence from a
particular patient, the user is first presented with clinical
context: differential diagnosis, phenotypes (collected at time of
order using standard ontology from SNOmed); additional clinical
information as available;
[0093] pedigree information; identity of the treating and/or
interpreting clinician. There are also areas to attach additional
information (for example, laboratory results, and/or clinical or
family history). The user may also look at technical summary to see
quality of the sequence data: for example, information about the
subject, a quality control report, and more detailed statistics
about the data, such as genome vs. exome coverage, or summary of
variants by various dimensions.
[0094] FIG. 2A is a screenshot 200 showing a display of genome
sequence data from a particular patient on the right side 210 in a
karyogram view. Each chromosome is numbered 211, and depicted
according to its banding pattern 212. Beside each chromosome are
markers that map variations determined by analysis of the test
sequence. The display on the left side 220 provides a list of
particular variations in a panel of genes of interest 221 which a
user has selected for analysis, with a summary assessment
underneath 222 as to whether it is pathogenic or benign. FIG. 2B is
a screenshot 230 comprising a cytogenomic view 240 of the entire
genome. Here, the data for each patient is compared with a sequence
standard drawn from CGI's database. FIG. 3A is an overview display
300 of the entire genome sequence of the patient sample in the form
of a circular or Circos.RTM.-style plot (M. Krzywinski et al.,
Genome Res. 19:1639-1645, 2009), with curved lines in the middle
301 depicting apparent interchromosomal junctions. FIG. 3B in this
example provides a detail 310 for Chromosome 4.
[0095] FIG. 4A is a display showing a chromosome view 400 for
Chromosome 1, selected for more detailed viewing by the user (for
example, by selecting a particular chromosome from the karyogram
display in FIG. 2A). The display in FIG. 4A shows the selected
chromosome 401 at the top, with markers and thumb controls 402
showing the region depicted in the graphics below. The linear plots
403, 404, 405, and 406 provide a map within the selected region of
the calculated copy number 403 (expected to be 2, except for
inter-regional duplication), the calculated allele specific copy
number 404 (expected to be 2 where homozygous and 1 where
heterozygous), the occurrence of small nucleotide variations 405,
and gene locations 406 (occurrence of open reading frames). From
this view, the user can zoom in and out using top thumb controls
402. A double tap zooms in, whereas a finger double tap zooms out.
By adjusting the position of the thumb controls 402, the user can
zoom towards a subregion of interest within the previous range.
Alternatively, the user may use the + and - buttons 407 on the
right to zoom the image from the center of the chromosomal
region.
[0096] FIG. 4B shows an alternative means by which a user may zoom
in to a particular gene. The screenshot 410 shows a keyboard 412
called up by the user, which can then be used to type the name of a
gene or location of interest into the search box 411. The system
then selects the appropriate chromosome and zooms into the
intrachromosomal region where the gene is located. The
magnification is selected by the system to accommodate the known
length of the gene (including introns, exons, and control
sequences), centering the gene between flanking sequence on each
side.
[0097] FIG. 4C is a display showing an expanded view 420 of a
narrower region of Chromosome 17 421 between the markers 422 on the
left. The user may reach this view by expanding a view of complete
Chromosome 17, zooming in on a particular gene, or by searching for
the gene by its name (TP53). Shown in this display are maps for two
overlapping genes: TP53 426 which is oriented left to right 426,
and a portion of WRAP53, which happens to partially overlap with
TP53, and is oriented right to left 427. Each gene is depicted as a
horizontal line with vertical lines on the 5' and 3' ends and a
directional indicator ("<" or ">") indicating the
orientation. Encoding regions for the unprocessed translation
product are indicated in block form at the corresponding location
within each gene.
[0098] FIG. 4D is a display 430 showing a further expanded view of
a particular gene on Chromosome 14 431. The user may reach this
level of expansion by expanding from a lower resolution as before,
or by tapping on the particular SNV in a display at lower
resolution (corresponding to the SNVs shown in strip 425 of FIG.
4C). Again, the copy number 433 and allele specific copy number 434
are shown. At this level of expansion, the genome sequence appears
as single bases 435. The test sequence and reference sequence are
the same, except where there is a variation: in this case, an SNV
437. The bottom-most graphic 436 is the encoded amino acid sequence
depicted using the one-letter code.
[0099] FIGS. 5A, 5B, 5C, and 5D show menus for selecting filters
that can be applied to the data, so as to simplify and focus the
variations shown within the selected region. Sequence variations of
particular interest are caused to be shown on the display, whereas
other variations or less interest or sequencing noise are filtered
out and do not appear.
[0100] FIG. 5A shows a menu of filters 500 to focus the analysis
towards certain variants of potential interest or importance. The
user may select specific genes 501 or specific chromosomal
locations 502 for the analysis. Small nucleotide variations (SNV)
can be filtered according to predicted pathogenicity, gene region,
impact on encoded amino acid sequence, SNV type (e.g., single
nucleotide polymorphisms, deletions, insertions, multi-nucleotide
substitutions), zygosity (homozygous or heterozygous), whether it
is found in a particular database such as dbSNP, to exclude
commonly found variants in healthy populations, and sequencing
quality filters (such as VQHIGH vs. VQLOW, and minimum read count).
Heterozygous variants marked as VQHIGH generally require at least
two high quality, well mapped reads per allele. Homozygous variants
marked as VQHIGH generally require at least seven reads. Variants
marked as VQLOW may have fewer reads supporting the call, and are
accompanied by a lower score indicating the lower confidence in the
call. Copy number variations (CNV) can be filtered according to
predicted pathogenicity, gene region, number of copies, repeat
category overlap, whether it is found in a particular external
database such as DGV, and sequencing quality filters, such as
ploidy quality score, and CNV type quality score.
[0101] FIG. 5B shows a menu 505 by which a user may assemble a gene
panel of particular interest. In this case, a panel of 15 genes is
selected. FIG. 5C shows a menu 510 where the user selects gene
regions by function 511 that should be included in the analysis.
Rather than showing the entire gene sequence, the user may select
to appear the encoding region (CDS), upstream transcription start
sites (TSS), introns, exon overlap at the 5' and/or 3' end, mRNA
splicing donor and acceptor sites and untranslated regions (UTR) at
the 5' and/or 3' end.
[0102] FIG. 5D shows a menu that allows the user to select SNVs
depending on their effect 521 on the encoded protein or its
expression: for example, no change, synonymous variations (silent
mutations that result in no change in encoded amino acid sequence),
nonsense variations (resulting in a premature stop codon), missense
variations (resulting in a chance in a single amino acid), a
misstart variation (resulting in misplacement of a start codon),
in-frame insertions or deletions (resulting in a small change in
amino acid sequence but leaving most of the protein intact),
frameshift insertions or deletions (causing downstream nucleic acid
sequence to be out of frame, thereby encoding an entirely different
amino acid sequence), splice site disruption (preventing normal
post-transcription processing), and unknown or unspecified
variations.
[0103] In this implementation, the user has an option to apply the
filters to either single nucleotide variations (SNVs), copy number
variations (CNVs), or both. For SNVs, the user may restrict the
analysis to variants where there is some information in the system
which indicates its pathogenicity. Other SNV filters include gene
region (what region the variant falls in, e.g. CDS, 5'UTR, 3'UTR),
the functional impact (e.g. synonymous, missense, nonsense, or
frameshift), SNV type (e.g., insertion, deletion, or substitution),
zygosity (homozygous or heterozygous), dbSNP(whether it's found in
dbSNP or not), minor Allele Freq for 1000 genomes to potentially
filter out commonly found variants in healthy populations, and
filters for sequence quality and minimum count. For CNVs, the user
may select filters for CNV: whether they should be included (for
example, CNVs greater than a given size), pathogenicity, copy
number state, repeat category overlap (e.g. DNA, LINE,
Low_Complexity, SINE, Satellite, SegDup, Self-chain,
Simple_Repeats, scRNA, tRNA, snRNA), whether it appears in the
Database of Genetic Variants (DGV), and quality filters (e.g.,
Ploidy Quality Score, CNV Type Quality Score). The Voyager system
allows the user may create their own filter having filtering
characteristics they desire.
[0104] FIG. 6 shows a display 600 with a summary of the analyzed
data characterized by various dimensions of interest. Each of the
genes 601 is tabulated against its predicted functional impact:
whether the SNV is synonymous 602, causes no change 603, has an
unknown or inconclusive result 604, constitutes a missense mutation
605, and the total 606. The numbers in the table are set up as
links If the user taps on a number, the variant list is filtered to
show the selected variants. The user can tap on a gene next to
Functional Impact Row to navigate back to previous screen,
conducting a search and presenting an expanded display for the
selected gene.
[0105] The system also provides links to external databases and
browsers such as UCSC or Ensembl so that the user can investigate
more details on gene structure or conservation information in a
gene region of interest, look up information in GeneReviews or do a
more global NCBI Search.
[0106] FIG. 7 shows a display 700 with a community assessment of a
particular sequence variation that has been detected by comparing
the test sequence to a reference sequence. At any time, the user
can create an assessment to capture research notes and other
evidence collected. When ready, the user may instruct the server to
share the assessment with other users of the system. As users
create and share assessments, Voyager then aggregates assessments
for a particular variant across cases and provide an average
community assessment for that variant. When that variant is seen
again in a particular case, users can use this aggregate assessment
as additional information in filtering, sorting and assessing
variants to fit the context of the specific case.
[0107] The bar chart 701 shows the distribution of community
assessments by rating. Underneath the chart, the user may access
written comments 702 from each of the contributors to the
assessment. As new assessments are created or comments are posted
on assessments, other members of community get notifications that
there is new activity. In the case when the user is not already
logged into Voyager, they would see iPad notifications. If users is
already in Voyager, they would see an indicator on a newsfeed icon
presented as part of the display. Where the user is in the context
of a specific case, the newsfeed is automatically filtered to
community activity for the given case. There is also a newsfeed on
the home page which shows activity across all cases.
[0108] FIG. 8 shows a screenshot 800 comprising a first window 801
summarizing the case with the number of variants, and indicating
when there are new comments. A second window 802 shows a map of
sequence variations shared with other users. A third window 804
provides information inputted from a first user, typically the user
who supplied the sequence being discussed. A fourth window 803 is
programmed and configured for exchange of commentary, where other
users of the system post reply information, which can initiate
further dialog from the first user and others.
VII. Flow of Information
[0109] FIG. 9 is a flow chart showing a system and process by which
a user may submit a sequence for processing and obtain a display
with the resulting analysis. After the user has securely logged in
to the system 921, they present a sequence from a test sample 922
stored in a secure sample sequence database 923. The user may then
select one or more particular reference sequences 924 from a
reference database 925 that matches the test sequence, or let the
server select reference sequence(s) that are appropriate. The test
and reference sequences are then aligned 926, and sequence
variations are identified 927.
[0110] The server then creates a map of variations 931, in which
differences between the test and reference sample(s) are located in
relation to known chromosomal markers. The server then compiles a
graphical display 932 of the variations, which is then presented to
the client 933. The display may be presented in a certain
preselected chromosomal region of interest. Alternatively, as
described in more detail below, the differences can be presented
according to the scope of the test sequence (for example, a
karyogram or chromosome view), from which the user may zoom in to
focus on the details of a particular region 934, or zoom back out.
The display is adjusted 935, and presented to the client 933
according to the request. As described below, the user may also
request one or more filters 936 which are used to select variations
between the test and reference samples 937 which are significant or
of particular interest. The variations remaining after the filters
are then used to create a map 931 and a graphical display 932 for
display to the client 933. The user may provide their own
annotation 941 for each variation which is incorporated 942 into
the display 932. In a multi-user system, other users may reply 943
to the first user's annotation. The reply information is then
incorporated into the display 942 either as a direct annotation or
as a linked comment.
VIII. Computer Implementation
[0111] As a general matter, computer systems referred to in the
context of this invention, their design, manufacture, control, and
programming may be conducted according to any suitable technology.
Any of the computer systems referred to in this disclosure can use
any suitable number of subsystems. A computer system may include a
single computer apparatus, where the subsystems can be the
components of the computer apparatus. Alternatively, a computer
system can include multiple computer apparatuses, each being a
subsystem, with internal components.
[0112] The subsystems shown in FIG. 10 are interconnected via a
system bus 1012. Additional subsystems such as a printer 1004,
keyboard 1008, fixed disk 1009, monitor 1006, which is coupled to
display adapter 1005, and others are shown. Peripherals and
input/output (I/O) devices, which couple to I/O controller 1001,
can be connected to the computer system by any number of means
known in the art, such as serial port 1007 (e.g. USB). For example,
serial port 1007 or external interface 1011 can be used to connect
computer system 1000 to a wide area network such as the Internet, a
mouse input device, or a scanner. The interconnection via system
bus 1012 allows the central processor 1003 to communicate with each
subsystem and to control the execution of instructions from system
memory 1002 or the fixed disk 1009, as well as the exchange of
information between subsystems. The system memory 1002 and/or the
fixed disk 1009 may embody a computer readable medium. Any of the
values mentioned herein can be output from one component to another
component and can be output to the user.
[0113] A computer system can include a plurality of the same
components or subsystems connected together by external interface
1011 or by an internal interface. Optionally, computer systems,
subsystem, or apparatuses can communicate over a network. In such
instances, one computer can be considered a client and another
computer a server, where each can be part of a same computer
system. A client and a server can each include multiple systems,
subsystems, or components.
[0114] Any of the embodiments of the invention can be implemented
in the form of control logic using hardware and/or using computer
software in a modular or integrated manner. Any of the software
components or functions in this disclosure may be implemented as
software code to be executed by a processor using any suitable
computer language such as, for example, Java, C++ or Perl using,
for example, conventional or object-oriented techniques. The
software code may be stored as a series of instructions or commands
on a computer readable medium for storage and/or transmission,
suitable media include random access memory (RAM), a read only
memory (ROM), a magnetic medium such as a hard-drive or a floppy
disk, or an optical medium such as a compact disk (CD) or DVD
(digital versatile disk), or flash memory. The computer readable
medium may be any combination of such storage or transmission
devices.
[0115] Computer programs may also be encoded and transmitted using
carrier signals adapted for transmission via wired, optical, and/or
wireless networks conforming to a variety of protocols, including
the Internet. As such, a computer readable medium may be created
using a data signal encoded with such programs. Computer readable
media encoded with the program code may be packaged with a
compatible device or provided separately from other devices (for
example, via Internet download). Any such computer readable medium
may reside on or within a single computer program product (such as
a hard drive, a CD, or an entire computer system), and may be
present on or within different computer program products within a
system or network. A computer system may include a monitor,
printer, or other suitable display for providing interaction and
display with the user.
[0116] Any of the methods referred to in this disclosure may be
totally or partially performed with a computer system including a
processor, which can be configured to perform the steps. Thus,
embodiments can be directed to computer systems configured to
perform the steps of any of the methods described here, potentially
with different components performing a respective steps or a
respective group of steps. Although presented as numbered steps,
steps of methods herein can be performed at a same time or in a
different order. Portions of these steps may be used with portions
of other steps from other methods. Also, all or portions of a step
may be optional. Additionally, any of the steps of any of the
methods can be performed with modules, circuits, or other means for
performing these steps.
[0117] The invention has been described and illustrated in this
disclosure with reference to particular embodiments for the benefit
and convenience of the reader. Discussion of various aspects of
products and methods does not limit manufacture or use of the
claimed invention except where expressly indicated.
[0118] The devices and methods of the invention may be substituted
and adapted for use in different contexts for different objectives
using different materials, elements, and steps without undue
experimentation, thus achieving any or all of the benefits of the
invention without departing from the scope of what is claimed.
[0119] In the United States of America and elsewhere as permitted
by law, each publication and patent document cited in this
disclosure is incorporated into the disclosure by reference in its
entirety for all purposes.
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